ABSTRACT T cell exhaustion is a differentiation state that is marked by the loss of effector function and increased expression of inhibitory receptors such as PD-1. Although virus-specific CD8 T cells are commonly considered as a homogeneous population that gradually become exhausted over time, recent research has clearly demonstrated that a CXCR5hi TCF-1hi subset is serving as a self-renewing progenitor population that can give rise to a more terminally exhausted CXCR5lo TCF-1lo subset. To better dissect the heterogeneity of ?exhausted? CD8 T cells, the lab applied single cell RNA-seq (scRNA-seq) to the chronic LCMV infection and identified three major subsets of virus-specific CD8 T cells that are phenotypically, functionally and transcriptionally distinct. Not only had the lab validated the existence of these subsets of T cells experimentally by flow cytometry, more advanced computational analyses were also performed to further predict their core transcriptional networks and developmental trajectories. Collectively, the findings reveal that a TCF-1hi progenitor subset can give rise to either a truly exhausted PD-1hi subset or a newly identified functional effector population that is named CX3CR1hi subset. This discovery laid a solid framework that allows testing of how extracellular signals and intrinsic genetic circuits regulate the formation and function of these three subsets of CD8 T cells. More importantly, it provides unprecedented opportunities to explore the possibility of generating more functional CX3CR1hi cells from TCF-1hi progenitors to overcome T cell exhaustion. This conceptual breakthrough is obviously applicable to control over chronic viral infection as well as cancer. Intriguingly, the preliminary study has also demonstrated that CD4 helper T cells, possibly through producing IL-21, are critical for TCF-1hi ? CX3CR1hi transition. This led to the hypothesis that inflammatory cytokines (such as CD4- derived IL-21) modulate the cellular, functional and transcriptional diversity of virus-specific CD8 T cells during chronic LCMV infection. Blocking antibodies, RNA interference and genetic deletion models will be used to further dissect how inflammatory cytokines and transcriptional networks regulate heterogeneity in T cell exhaustion. Furthermore, the lab proposes to redirect CD8 T cell differentiation away from ?exhaustion? by providing additional ?CD4 help?, either alone or in combination with PD-1 blockade. The lab will test if providing IL-21 producing CD4 T cells through adoptive transfer could drive TCF-1hi progenitor cell differentiation into functional CX3CR1hi effector cells. Overall, knowledge gained from this research will provide mechanistic insights into how to redirect the formation of functional effector T cells and simultaneously limit T cell exhaustion for improved viral control over chronic infection.